Department of Orthopedics, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030, China.
Department of Orthopedic Surgery, Second Affiliated Hospital, School of Medicine, Zhejiang University, Hangzhou, China.
Biomed Res Int. 2018 May 30;2018:3936257. doi: 10.1155/2018/3936257. eCollection 2018.
FNDC4 acts as an anti-inflammatory factor on macrophages and improves mouse model of induced colitis. Considering osteoclast formation is characterized by the activation of inflammation-related pathways, we thus speculated that FNDC4 may play a pivotal role in this process. RT-qPCR analysis was performed to confirm the expression of osteoclast formation related genes in primary murine bone marrow macrophages (BMMs). RANKL-treated BMMs were cultured with FNDC4 to evaluate the effect of FNDC4 on osteoclast differentiation. TRAP staining and bone resorption pits assay were used to assess osteoclast formation and bone resorption, respectively. Luciferase assay and western blotting analysis were conducted to determine whether FNDC4 inhibited osteoclast formation via NF-B signaling in RAW 264.7 cells. Furthermore, to identify gene signatures in FNDC4-treated BMMs and to use these to elucidate the underlying molecular mechanisms during osteoclast formation, we adopted a bioinformatics approach by downloading the GSE76172 gene expression profiling dataset from the Gene Expression Omnibus (GEO) database. FNDC4 inhibited RANKL-induced osteoclastogenesis and mature osteoclast resorptive function in a dose-dependent manner. Results of NF-B luciferase assay suggested that FNDC4 could significantly suppress the RANKL-induced NF-B transcriptional activity. Based on the protein-protein interaction network, CXCL10 was identified as the differentially expressed gene with the highest connectivity degree (degree = 23); it was drastically downregulated in the presence of FNDC4, but supplementation of CXCL10 (10 ng/mL) partially ameliorated the FNDC4-induced inhibition of osteoclast formation. Taken together, we speculated that FNDC4 could suppress osteoclast formation via NF-B pathway and downregulation of CXCL10.
FNDC4 作为一种抗炎因子作用于巨噬细胞，并改善诱导结肠炎的小鼠模型。鉴于破骨细胞的形成以炎症相关途径的激活为特征，我们因此推测 FNDC4 可能在这个过程中发挥关键作用。我们进行了 RT-qPCR 分析以确认原代小鼠骨髓巨噬细胞（BMM）中破骨细胞形成相关基因的表达。用 FNDC4 培养 RANKL 处理的 BMM 以评估 FNDC4 对破骨细胞分化的影响。TRAP 染色和骨吸收陷窝试验分别用于评估破骨细胞形成和骨吸收。萤光素酶测定和 Western blot 分析用于确定 FNDC4 是否通过 RAW 264.7 细胞中的 NF-B 信号通路抑制破骨细胞形成。此外，为了鉴定 FNDC4 处理的 BMM 中的基因特征，并利用这些特征阐明破骨细胞形成过程中的潜在分子机制，我们采用了生物信息学方法，从基因表达综合数据库（GEO）下载了 GSE76172 基因表达谱数据集。FNDC4 以剂量依赖性方式抑制 RANKL 诱导的破骨细胞生成和成熟破骨细胞的吞噬功能。NF-B 萤光素酶测定结果表明 FNDC4 可以显著抑制 RANKL 诱导的 NF-B 转录活性。基于蛋白质-蛋白质相互作用网络，CXCL10 被鉴定为具有最高连接度（度=23）的差异表达基因；FNDC4 存在时其表达明显下调，但补充 CXCL10（10ng/mL）部分缓解了 FNDC4 诱导的破骨细胞形成抑制。总之，我们推测 FNDC4 可以通过 NF-B 通路和下调 CXCL10 来抑制破骨细胞形成。
